In vivo acoustic patterning of endothelial cells for tissue vascularization.

Strategies to fabricate microvascular networks that structurally and functionally mimic native microvessels are needed to address a host of clinical conditions associated with tissue ischemia. The objective of this work was to advance a novel ultrasound technology to fabricate complex, functional microvascular networks directly in vivo. Acoustic patterning utilizes forces within an ultrasound standing wave field (USWF) to organize cells or microparticles volumetrically into defined geometric assemblies.

Receptor-binding domain of SARS-CoV-2 is a functional αv-integrin agonist.

Among the novel mutations distinguishing SARS-CoV-2 from similar coronaviruses is a K403R substitution in the receptor-binding domain (RBD) of the viral spike (S) protein within its S1 region. This amino acid substitution occurs near the angiotensin-converting enzyme 2-binding interface and gives rise to a canonical RGD adhesion motif that is often found in native extracellular matrix proteins, including fibronectin. Here, the ability of recombinant S1-RBD to bind to cell surface integrins and trigger downstream signaling pathways was assessed and compared with RGD-containing, integrin-binding fragments of fibronectin.

Receptor binding domain of SARS-CoV-2 is a functional αv-integrin agonist.

Among the novel mutations distinguishing SARS-CoV-2 from similar respiratory coronaviruses is a K403R substitution in the receptor-binding domain (RBD) of the viral spike (S) protein within its S1 region. This amino acid substitution occurs near the angiotensin-converting enzyme 2 (ACE2)-binding interface and gives rise to a canonical RGD adhesion motif that is often found in native extracellular matrix proteins, including fibronectin. In the present study, the ability of recombinant S1-RBD to bind to cell surface integrins and trigger downstream signaling pathways was assessed and compared to RGD-containing, integrin-binding fragments of fibronectin.

Using Acoustic Fields to Fabricate ECM-Based Biomaterials for Regenerative Medicine Applications.

Ultrasound is emerging as a promising tool for both characterizing and fabricating engineered biomaterials. Ultrasound-based technologies offer a diverse toolbox with outstanding capacity for optimization and customization within a variety of therapeutic contexts, including improved extracellular matrix-based materials for regenerative medicine applications. Non-invasive ultrasound fabrication tools include the use of thermal and mechanical effects of acoustic waves to modify the structure and function of extracellular matrix scaffolds both directly, and indirectly via biochemical and cellular mediators.

Acoustic Fabrication of Collagen-Fibronectin Composite Gels Accelerates Microtissue Formation.

Ultrasound can influence biological systems through several distinct acoustic mechanisms that can be manipulated by varying reaction conditions and acoustic exposure parameters. We recently reported a new ultrasound-based fabrication technology that exploits the ability of ultrasound to generate localized mechanical forces and thermal effects to control collagen fiber microstructure non-invasively. Exposing solutions of type I collagen to ultrasound during the period of microfibril assembly produced changes in collagen fiber structure and alignment, and increased the biological activity of the resultant collagen hydrogels.

Time- and Dose-Dependent Effects of Pulsed Ultrasound on Dermal Repair in Diabetic Mice.

Chronic wounds, including diabetic, leg and pressure ulcers, impose a significant health care burden worldwide. Some evidence indicates that ultrasound can enhance soft tissue repair. However, therapeutic responses vary among individuals, thereby limiting clinical translation.

CD4 T Cell Interstitial Migration Controlled by Fibronectin in the Inflamed Skin.

The extracellular matrix (ECM) is extensively remodeled during inflammation providing essential guidance cues for immune cell migration and signals for cell activation and survival. There is increasing interest in the therapeutic targeting of ECM to mitigate chronic inflammatory diseases and enhance access to the tumor microenvironment. T cells utilize the ECM as a scaffold for interstitial migration, dependent on T cell expression of matrix-binding integrins αβ/αβ and tissue display of the respective RGD-containing ligands.

Non-invasive acoustic fabrication methods to enhance collagen hydrogel bioactivity.

Much attention has focused recently on utilizing components of the extracellular matrix (ECM) as natural building blocks for a variety of tissue engineering applications and regenerative medicine therapies. Consequently, new fabrication methods are being sought to enable molecular control over the structural characteristics of ECM molecules in order to improve their biological function. Exposing soluble collagen to acoustic forces associated with ultrasound propagation produces localized variations in collagen microfiber organization that in turn, promote cell behaviors essential for tissue regeneration, including cell migration and matrix remodeling.

A Matricryptic Conformation of the Integrin-Binding Domain of Fibronectin Regulates Platelet-Derived Growth Factor-Induced Intracellular Calcium Release.

Platelet-derived growth factor (PDGF) signaling is dysregulated in a wide variety of diseases, making PDGF an attractive therapeutic target. However, PDGF also affects numerous signaling cascades essential for tissue homeostasis, limiting the development of PDGF-based therapies that lack adverse side-effects. Recent studies showed that fibroblast-mediated assembly of extracellular matrix (ECM) fibronectin fibrils attenuates PDGF-induced intracellular calcium release by selectively inhibiting phosphoinositol 3-kinase (PI3K) activation while leaving other PDGF-mediated signaling cascades intact.

Assembly of fibronectin fibrils selectively attenuates platelet-derived growth factor-induced intracellular calcium release in fibroblasts.

Cellular responses to platelet-derived growth factor (PDGF) are altered in a variety of pathological conditions, including cancers, fibroses, and vascular diseases, making PDGF-induced signaling pathways important therapeutic targets. The limited success of therapies designed to impact PDGF pathways may be overcome with a clearer understanding of how cells integrate signals from PDGF and the extracellular matrix (ECM). Here, we assessed the effects of fibronectin matrix assembly on the responsiveness of mesenchymal cells to PDGF.

A Small Chimeric Fibronectin Fragment Accelerates Dermal Wound Repair in Diabetic Mice.

During wound repair, soluble fibronectin is converted into biologically active, insoluble fibrils via a cell-mediated process. This fibrillar, extracellular matrix (ECM) form of fibronectin stimulates cell processes critical to tissue repair. Nonhealing wounds show reduced levels of ECM fibronectin fibrils.

Ultrasound patterning technologies for studying vascular morphogenesis in 3D.

Investigations in this report demonstrate the versatility of ultrasound-based patterning and imaging technologies for studying determinants of vascular morphogenesis in 3D environments. Forces associated with ultrasound standing wave fields (USWFs) were employed to non-invasively and volumetrically pattern endothelial cells within 3D collagen hydrogels. Patterned hydrogels were composed of parallel bands of endothelial cells located at nodal regions of the USWF and spaced at intervals equal to one half wavelength of the incident sound field.

Extracellular matrix fibronectin mediates an endothelial cell response to shear stress via the heparin-binding, matricryptic RWRPK sequence of FNIII1H.

Endothelial cells (EC) respond to mechanical forces such as shear stress in a variety of ways, one of which is cytoskeletal realignment in the direction of flow. Our earlier studies implicated the extracellular matrix protein fibronectin in mechanosensory signaling to ECs in intact arterioles, via a signaling pathway dependent on the heparin-binding region of the first type III repeat of fibrillar fibronectin (FNIII1H). Here we test the hypothesis that FNIII1H is required for EC stress fiber realignment under flow.

Cooperative effects of fibronectin matrix assembly and initial cell-substrate adhesion strength in cellular self-assembly.

Unlabelled: The cell-dependent polymerization of intercellular fibronectin fibrils can stimulate cells to self-assemble into multicellular structures. The local physical cues that support fibronectin-mediated cellular self-assembly are largely unknown. Here, fibronectin matrix analogs were used as synthetic adhesive substrates to model cell-matrix fibronectin fibrils having different integrin-binding specificity, affinity, and/or density.

Extracellular matrix fibronectin initiates endothelium-dependent arteriolar dilatation via the heparin-binding, matricryptic RWRPK sequence of the first type III repeat of fibrillar fibronectin.

The local arteriolar dilatation produced by contraction of skeletal muscle is dependent upon multiple signalling mechanisms. In addition to the many metabolic signals that mediate this vasodilatation, we show here that the extracellular matrix protein fibronectin also contributes to the response. This vasodilatory signal requires the heparin-binding matricryptic RWRPK sequence in the first type III repeat of fibrillar fibronectin.

Quantitative Ultrasound for Nondestructive Characterization of Engineered Tissues and Biomaterials.

Non-invasive, non-destructive technologies for imaging and quantitatively monitoring the development of artificial tissues are critical for the advancement of tissue engineering. Current standard techniques for evaluating engineered tissues, including histology, biochemical assays and mechanical testing, are destructive approaches. Ultrasound is emerging as a valuable tool for imaging and quantitatively monitoring the properties of engineered tissues and biomaterials longitudinally during fabrication and post-implantation.

Biological Effects of Low-Frequency Shear Strain: Physical Descriptors.

Biological effects of megahertz-frequency diagnostic ultrasound are thoroughly monitored by professional societies throughout the world. A corresponding, thorough, quantitative evaluation of the archival literature on the biological effects of low-frequency vibration is needed. Biological effects, of course, are related directly to what those exposures do physically to the tissue-specifically, to the shear strains that those sources produce in the tissues.

Scholte wave generation during single tracking location shear wave elasticity imaging of engineered tissues.

The physical environment of engineered tissues can influence cellular functions that are important for tissue regeneration. Thus, there is a critical need for noninvasive technologies capable of monitoring mechanical properties of engineered tissues during fabrication and development. This work investigates the feasibility of using single tracking location shear wave elasticity imaging (STL-SWEI) for quantifying the shear moduli of tissue-mimicking phantoms and engineered tissues in tissue engineering environments.

Therapeutic Applications of Extracellular Matrix.

Chronic and hard-to-heal wounds are a tremendous burden on our healthcare system and impair the quality of life for millions of people. An emerging focus of regenerative medicine is the development of natural biomaterials that can stimulate tissue formation or repair by recreating the functional and structural properties of proteins and polysaccharides found within the extracellular matrix (ECM). Promising new developments include the fabrication of novel ECM-based biologics to selectively deliver drugs or growth factors to wounds; new classes of bioactive tissue sealants, scaffolds, and hydrogels; as well as inductive wound dressings derived from decellularized tissues.

Noninvasive Quantitative Imaging of Collagen Microstructure in Three-Dimensional Hydrogels Using High-Frequency Ultrasound.

Collagen I is widely used as a natural component of biomaterials for both tissue engineering and regenerative medicine applications. The physical and biological properties of fibrillar collagens are strongly tied to variations in collagen fiber microstructure. The goal of this study was to develop the use of high-frequency quantitative ultrasound to assess collagen microstructure within three-dimensional (3D) hydrogels noninvasively and nondestructively.

Ultrasound technologies for biomaterials fabrication and imaging.

Ultrasound is emerging as a powerful tool for developing biomaterials for regenerative medicine. Ultrasound technologies are finding wide-ranging, innovative applications for controlling the fabrication of bioengineered scaffolds, as well as for imaging and quantitatively monitoring the properties of engineered constructs both during fabrication processes and post-implantation. This review provides an overview of the biomedical applications of ultrasound for imaging and therapy, a tutorial of the physical mechanisms through which ultrasound can interact with biomaterials, and examples of how ultrasound technologies are being developed and applied for biomaterials fabrication processes, non-invasive imaging, and quantitative characterization of bioengineered scaffolds in vitro and in vivo.

Estimating cell concentration in three-dimensional engineered tissues using high frequency quantitative ultrasound.

Histology and biochemical assays are standard techniques for estimating cell concentration in engineered tissues. However, these techniques are destructive and cannot be used for longitudinal monitoring of engineered tissues during fabrication processes. The goal of this study was to develop high-frequency quantitative ultrasound techniques to nondestructively estimate cell concentration in three-dimensional (3-D) engineered tissue constructs.

Opposing effects of collagen I and vitronectin on fibronectin fibril structure and function.

Extracellular matrix fibronectin fibrils serve as passive structural supports for the organization of cells into tissues, yet can also actively stimulate a variety of cell and tissue functions, including cell proliferation. Factors that control and coordinate the functional activities of fibronectin fibrils are not known. Here, we compared effects of cell adhesion to vitronectin versus type I collagen on the assembly of and response to, extracellular matrix fibronectin fibrils.

Regional fibronectin and collagen fibril co-assembly directs cell proliferation and microtissue morphology.

The extracellular matrix protein, fibronectin stimulates cells to self-assemble into three-dimensional multicellular structures by a mechanism that requires the cell-dependent conversion of soluble fibronectin molecules into insoluble fibrils. Fibronectin also binds to collagen type I and mediates the co-assembly of collagen fibrils into the extracellular matrix. Here, the role of collagen-fibronectin binding in fibronectin-induced cellular self-assembly was investigated using fibronectin-null fibroblasts in an in vitro model of tissue formation.

Controlling collagen fiber microstructure in three-dimensional hydrogels using ultrasound.

Type I collagen is the primary fibrillar component of the extracellular matrix, and functional properties of collagen arise from variations in fiber structure. This study investigated the ability of ultrasound to control collagen microstructure during hydrogel fabrication. Under appropriate conditions, ultrasound exposure of type I collagen during polymerization altered fiber microstructure.